Theoretical Study On Substrate Selectivity Of Cytochrome P450 2D6 And 1A2

Cytochrome P450(CYP)is a large heme protein family,belonging to a monooxygenase,named for its specific absorption peak at 450 nm.Most members of CYP are membrane proteins,which are distributed in the endoplasmic reticulum and mitochondrial inner membrane of most tissues and cells in human body.Scientists have found that CYPs are closely related to the metabolism of various drugs in the liver.It can catalyze most drugs and convert them from hydrophobic type to hydrophilic type,which is easier to metabolize.However,due to the large individual diversity of CYPs and the complex catalytic mechanism,it is not an easy job to carry out the related studies,so the interaction between CYPs and drug molecules has been a hot topic in the research field for many years.In this paper,multiple templates approach was used to construct the all-sequence structure of CYP1A2 and CYP2D6,two enzymes that contribute significantly to drug metabolism in the CYPs family,and the interaction between CYP1A2 and CYP2D6 and drug molecules was detailedly studied by means of molecular dynamics simulation and quantum mechanics calculation.The main results are as follows:1.The Regioselectivity of the Interaction Between Dextromethorphan and CYP2D6Nearly 25% of drugs sold on the market are metabolized by CYP2D6.During the decades,scientists have made remarkable achievements in the chemical mechanism of the reaction of substrates catalyzed by CYP2D6 and the influence of amino acid mutation of CYP2D6 on the interaction with substrates.However,the causes of the regioselectivity for CYP2D6-catalyzed reactions and the influence of substrate concentration on the interaction between CYP2D6 and substrates still remain unclear.Therefore,dextromethorphan(DM),a classic metabolic substrate of CYP2D6,was selected as the molecule probe.Four research systems with different substrate orientations and substrate concentrations were constructed.The reasons for the regioselectivity of CYP2D6-catalyzed DM-demethylation reaction and the influence of DM concentration on the interaction were investigated based on their molecular interactions.According to our results,we found a new binding mode between CYP2D6 and DM,in which two key residues of CYP2D6,Asp301 and Glu216,can simultaneously form water-bridge hydrogen bonds with DM molecule to help it stabilize near the catalytic pocket of CYP2D6.In addition,we also found that when substrate concentration increased,the volume of the catalytic pocket of CYP2D6 would decrease,thus reducing the activity of CYP2D6-catalyzed O-demethylation of DM,but increasing the activity of CYP2D6-catalyzed N-demethylation of DM.These results contribute to a better understanding of the interactions between CYP2D6 and substrates,and provide theoretical guidance for experimental studies and drug design related to CYP2D6 to some extent.2.The Substrate Inhibition Mechanism of CYP2D6 Metabolizing DextromethorphanIn the previous study,we found that the multiple substrate binding event can reduce the activity of CYP2D6 metabolizing substrates,but the corresponding theoretical mechanism has rarely been reported,which aroused our interest in exploring the mechanism.Here,we combined molecular dynamics simulation with QM calculation to explore the mechanism of how does multiple substrate binding lead to the substrate inhibition of CYP2D6 metabolizing DM.According to our results,three gate residues(Arg221,Val374,and Phe483)for the catalytic pocket were determined.We also found that when the second substrate molecule binds to the site in channel 2b,the secondary structure near the catalytic pocket of CYP2D6 has a relative movement trend,which compresses the volume of the catalytic pocket of CYP2D6,and also affects the binding mode of DM in the catalytic pocket of CYP2D6 through steric effect.These changes decrease the stability of DM binding in the CYP2D6 catalytic pocket.In addition,the combination of the second substrate molecule also increases the activation energy of the substrate reaction catalyzed by CYP2D6,thus reducing the ability of the substrate reaction catalyzed by CYP2D6.Our findings would help to understand the substrate inhibition of CYP2D6 catalyzing the DM,and enrich the knowledge of the drug-drug interactions for the cytochrome P450 superfamily.3.A Theoretical Explanation for The Regioselectivity of CYP1A2-CatalyzedCaffeine Reaction Based on Molecular InteractionsCaffeine is a very common food ingredient,found in the daily diet of 90% of adults around the world.Caffeine can help people reduce fatigue,stay awake and increase productivity.It is even used before many sporting events by athletes in order to achieve better results.In human body,caffeine is mainly metabolized by CYP1A2,it has four metabolic sites(N1,N3,N7 and C8)and 80% of caffeine metabolism occurs at the N3 site.So far,there are few theoretical studies on the interaction between caffeine and CYP1A2,and the active binding modes between caffeine and CYP1A2 are still unclear.Therefore,in this part,we constructed the all-sequence structure of CYP1A2 by means of the multiple templates approach and performed molecular dynamics simulations to explore the binding modes between caffeine and CYP1A2.We try to explore the reasons why the N3 site of caffeine becomes the main metabolic site of CYP1A2 based on their molecular interactions.According to our results,four active binding modes between CYP1A2 and caffeine that correspond to the four metabolic sites of caffeine are discovered,among which the active binding mode of N3 site has the shortest distance between the metabolic sites and is the most stable one.This may be the reason why the N3 site of caffeine is the primary catalytic site for CYP1A2.In addition,a pre-reaction state for the CYP1A2-catalyzed reaction at caffeine’s N3 site is identified.These findings could enhance the knowledge of the interactions between CYP1A2 and caffeine,and help better understand how caffeine is metabolized in the body.